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Periodic Report Summary - BCELLSMICETOMEN (Translational study on the antigen presenting properties of human vs mouse B cell subpopulations)

The main scientific objectives of the project are to study comparatively the antigen presenting properties of mouse and human B cell subpopulations in view of their possible use as vectors of antigens in cell base cancer vaccines. To this end, the team had to set up a new laboratory, implementing a set of techniques including magnetic and flow cytometric sorting of lymphocyte subsets, primary immune response model based on two way mixed lymphocyte reaction, non-radioactive assay of T cell proliferation and cytokine production assay both based on intracellular staining and flow cytometry as well as bead based flow cytometric cytokine array. These techniques would then be applied reciprocally in the human system during the second period to collect a matching set of data that will permit the comparison of the antigen presenting properties of the studied B cell subsets. The main results achieved during the period are:

1. confirmation of a better stimulation to proliferation and type response differentiation by marginal zone as compared to follicular B cells as well as confirmation of the suppressive action of T2 (CD21hiCD23hiCD93+) B cells on T cell stimulation by allogeneic follicular or marginal zone B cells;
2. observation of opposite effect of T1 B cells (CD21loCD23loCD93+) on T cell stimulation by allogeneic follicular or marginal zone B cells - suppressive for the follicular B cells but augmenting for the marginal zone B cells;
3. observation of stimulation of T cells differentiation towards a population producing less interferon gamma (IFNgamma) and more interleukin 10 (IL-10) by CD21loCD23loCD93+ B cells.

These results will be further confirmed by a more precise sorting of the B cell subpopulations, extensive cytokine profile using a cytokine bead array and further elucidating the possible contribution of plasma cell in the CD21loCD23loCD93+ population. In the next stage, similar techniques will be applied to the main human B cell subsets to compare their stimulatory capacity for primary and secondary immune response. If the funding permits, at the final stage a mouse tumour model will be applied to test the capacity of the different B cell subsets, with or without further stimulation, to induce cytotoxic antitumour responses to particular tumour antigens in view of designing an optimised approach for B cell based cancer vaccine. As the efficient targeting of the antigen to the endosomal compartment facilitating presentation in MHC class II is crucial and the B cell population will be of diverse B cell receptor specificities, targeting techniques (e.g. based on conjugation to anti-IgM Fab antibody fragments) will be tested. Finally, an attempt will be made to perform transcriptomic comparison between mixed lymphocyte cultures of T cells stimulated with the best candidate (in terms of being practically sortable and sufficient in numbers) among the B cell subsets and its putative human counterpart. This last experiment will depend on collaboration with Winthrop P. Rockefeller Cancer Institute in Little Rock, Arkansas (AR).

The overall impact of these studies is related to the development new alternatives to the dendritic cell based vaccines aimed at improving the cellular yields and the phenotypic profile of the stimulated T cells.

Reported by

26, Acad. Georgi Bonchev St.
1113 SOFIA
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